Neutralization of Botulinum Neurotoxin by a Human Monoclonal Antibody Specific for the Catalytic Light Chain

Adekar, Sharad P.; Takahashi, Tsuyoshi; Jones, R. Mark; Al-Saleem, Fetweh H.; Ancharski, Denise M.; Root, Michael J.; Kapadnis, B. P.; Simpson, Lance L.; Dessain, Scott

doi:10.1371/journal.pone.0003023

Cited by 61 publications

(85 citation statements)

References 32 publications

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“…In the other scenario, the nLC would be in contact with the HA3 trimer, whereas the BoNT LC was at the distal end of the complex. To distinguish between these two possibilities, we labeled the PC with a BoNT/A1 LC-specific monoclonal antibody (4LCA) (25). We observed consistent and efficient labeling of the distal end of the ovoid body ( Fig.…”

Section: Resultsmentioning

confidence: 92%

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Molecular assembly of botulinum neurotoxin progenitor complexes

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et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Botulinum neurotoxin (BoNT) is produced by Clostridium botulinum and associates with nontoxic neurotoxin-associated proteins to form high-molecular weight progenitor complexes (PCs). The PCs are required for the oral toxicity of BoNT in the context of food-borne botulism and are thought to protect BoNT from destruction in the gastrointestinal tract and aid in absorption from the gut lumen. The PC can differ in size and protein content depending on the C. botulinum strain. The oral toxicity of the BoNT PC increases as the size of the PC increases, but the molecular architecture of these large complexes and how they contribute to BoNT toxicity have not been elucidated. We have generated 2D images of PCs from strains producing BoNT serotypes A1, B, and E using negative stain electron microscopy and single-particle averaging. The BoNT/A1 and BoNT/B PCs were observed as ovoid-shaped bodies with three appendages, whereas the BoNT/E PC was observed as an ovoid body. Both the BoNT/A1 and BoNT/B PCs showed significant flexibility, and the BoNT/B PC was documented as a heterogeneous population of assembly/disassembly intermediates. We have also determined 3D structures for each serotype using the random conical tilt approach. Crystal structures of the individual proteins were placed into the BoNT/A1 and BoNT/B PC electron density maps to generate unique detailed models of the BoNT PCs. The structures highlight an effective platform that can be engineered for the development of mucosal vaccines and the intestinal absorption of oral biologics.hemagglutinin | nontoxic nonhemagglutinin | sialic acid

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Section: Resultsmentioning

confidence: 92%

“…The 4LCA antibody was purified as previously described (25). For labeling, the BoNT/A1 PC and 4LCA proteins were mixed in a ∼1:10 molar ratio and incubated at room temperature for 3 h before grid preparation.…”

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confidence: 99%

Molecular assembly of botulinum neurotoxin progenitor complexes

Benefield

Dessain

Shine

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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“…While polyclonal antitoxin products are available to treat human botulism, it is widely accepted that improved antitoxin agents are badly needed (27). One approach showing promise is to replace polyclonal antisera with monoclonal antibodies (MAbs) (1,2,4,9,17,20). This approach has been applied successfully in mice, using anti-BoNT MAbs (17).…”

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Efficient Serum Clearance of Botulinum Neurotoxin Achieved Using a Pool of Small Antitoxin Binding Agents

et al. 2010

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Antitoxins for botulinum neurotoxins (BoNTs) and other toxins are needed that can be produced economically with improved safety and shelf-life properties compared to conventional therapeutics with large-animal antisera. Here we show that protection from BoNT lethality and rapid BoNT clearance through the liver can be elicited in mice by administration of a pool of epitope-tagged small protein binding agents together with a single anti-tag monoclonal antibody (MAb). The protein binding agents used in this study were single-chain Fv domains (scFvs) with high affinity for BoNT serotype A (BoNT/A). The addition of increasing numbers of differently tagged scFvs synergistically increased the level of protection against BoNT/A. It was not necessary that any of the BoNT/A binding agents possess toxin-neutralizing activity. Mice were protected from a dose equivalent to 1,000 to 10,000 50% lethal doses (LD 50 ) of BoNT/A when given three or four different anti-BoNT scFvs, each fused to an E-tag peptide, and an anti-E-tag IgG1 MAb. Toxin protection was enhanced when an scFv contained two copies of the E tag. Pharmacokinetic studies demonstrated that BoNT/A was rapidly cleared from the sera of mice given a pool of anti-BoNT/A scFvs and an anti-tag MAb but not from the sera of mice given scFvs alone or anti-tag MAb alone. The scFv pool and anti-tag MAb protected mice from lethality when administered up to 2 h following exposure of mice to a dose equivalent to 10 LD 50 of BoNT/A. These results suggest that it will be possible to rapidly and economically develop and produce therapeutic antitoxins consisting of pools of tagged binding agents that are administered with a single, stockpiled anti-tag MAb.

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“…In addition, neutralizing antibodies that target the light chain of BoNT/A through another mechanism, potentially by preventing translocation of the LC but not endocytosis of the holotoxin, have been found (54)(55)(56). Numerous epitopes on both the N-and C-terminal regions of HCR/A have been described, and neutralizing activity is absent from antibodies that target the N terminus, distal to the GBP (17,30,57,58).…”

Section: Discussionmentioning

confidence: 99%